Process for the treating of wood chips with fungi to enhance enzymatic hydrolysis of the resinous components



Dec. 30. 1969 T. NILSSON E AL PROCESS FOR THE TREATING OF WOOD CHIPS WITH FUNGI TO ENCHANCE ENZYMATIC HYDROLYSIS OF THE RESINOUS COMPONENTS Filed July 19, 1966 United States Patent 0.

3,486,969 PROCESS FOR THE TREATING F WOOD CHIPS WITH FUNGI TO ENHANCE ENZYMATIC HY- DROLYSIS OF THE RESINOUS COMPONENTS Thomas Nilsson, Stockholm, and Anders Assarsson, Ornskoldsvik, Sweden, assignors to M0 och Domsjo Aktiebolag, Ornskoldsvik, Sweden, a corporation of Sweden Filed July 19, 1966, Ser. No. 566,309 Claims priority, application Sweden, July 20, 1965, 9,591/ 65 Int. Cl. D21]: 1/02 US. Cl. 162-72 4 Claims ABSTRACT OF THE DISCLOSURE A dispersion of selected fungi is added to wood chips which are stored prior to further processing such as by chemical digestion to form pulp. The chips are subjected to enzymatic hydrolysis induced by the fungi while minimizing microbial \action on cellulose and hemicellulose in the wood.

The present invention relates to a process for storing wood chips whereby said chips are stacked in piles, and to an arrangement for carrying out the method.

When preparing cellulose pulp the resin components present in the wood give rise to certain problems being difiicult and costly to remove, causing disturbances in production and impairing the qu'ality'of the finished product. Resin from, predominantly, hardwood and softwood comprises hydrocarbons, waxes (esters between fatty acids and higher alcohols), glycerides and higher alcohols. Furthermore, present in softwood are diterpene acids (resin acids) and diterpene aldehydes. One method of facilitating the removal of these compounds is to store the barked or unbarked wood as round logs, on land or in water. When storing in water an enzymatical hydrolysis of the glycerides of the resin takes place. When storing on dry ground an autoxidation of saturated resin components takes place, in addition to the hydrolysis of glycerides, when the wood dries out. Both enzymatical hydrolysis and autoxidation facilitate removal of the resin in later pulp manufacturing processes. Round-timber storage, however, takes a long time and, furthermore, gives rise to a certain decomposition of cellulose and hemicellulose in the wood. To reduce these disadvantages investigations have been made as to the possibility of storing wood in the form of piles of chips, in which an enzymatical hydrolysis, autoxidation and a biological decomposition of all resin components takes place. However, when adopting these means :a biological breakdown of cellulose and hemicellulose also takes place, which causes the loss of valuable wood substances. Decomposition of resin has been discovered to give rise to resin losses which can substantially be attributed to the fatty acids released on the enzymatical hydrolysis of fats and waxes, which have thus been sujected to biological decomposition reactions. The enzymatical hydrolysis of the waxes is an advantage which is not obtained in any other method of storing wood. The waxes are extraordinarily difficult to remove during the pulping process due to the fact that their ester linkages are sterically hindered which make saponification almost impossible. This is illustrated by the fact that waxes in wood which is digested according to the sulphate method to 90% 'ice survive this treatment without being saponified and that the alkali treatment to which the pulp is subjected in the bleaching plant influences the ester linkages of the waxes to a still less degree. The waxes therefore enter into the finished pulp product and impair its quality. The present invention is built on the hypothesis that microorganisms present in stacks of 'wood chips have a decisive significance for the aging of the resins and the decomposition of cellulose and hemicellulose. The method according to the invention is characterized in that during the process of stacking the piles of wood chips, said chips are supplied with a culture of one or more microorganisms which break down the resin components present in the chips but do not decompose the cellulose, said microorganisms, moreover, preferably having an inhibiting effeet on other cellulose and hemicellulose decomposing microorganisms normally appearing in the pile of wood chips.

By these means it is possible to obtain an appreciably shortened storage time. in comparison with the storage of wood as logs whilst at the same time obtaining an extraordinarilly good resin ageing without the simultaneous decomposition of cellulose or hemicellulose, and to remove the waxes so disadvantageous to the pulp processes. The microoganisms may also possibly be added together with a nutritional salt solution. The inhibition of cellulose or hemicellulose decomposing microorganisms may in principle be effected in three different ways. Firstly it is possible to elect to add such a microorganism which, at the same time as it decomposes the resin constituents, inhibits the cellulose and hemicellulose decomposing microorganisms by generating such a high temperature in the pile of wood chips that the latter microorganisms are prevented from propagating. In this respect the said microorganisms can only withstand a temperature of, at the most approximately 50 C. Another method is to add a resin decomposing micro-organism which by an anti-biotic effect prevents the cellulose and hemicellulose decomposing micro-organisms from propagating. A third method is, at the same time as the specific resin decomposing micro-organism is added, to add a substance which has an inhibiting effect on the cellulose and hemicellulose decomposing micro-organisms, e.g. nickel sulphate. Naturally it is also possible to use combinations of these three methods of inhibiting cellulose and hemi-cellulose decomposing micro-organisms.

Suitable micro-organisms which can be used according to the present invention are fungi, which completely or partly lack the ability of producing active celluloses on wood but which can produce esterases such as lipases, sterolases, sterolesterases, fatty alcoholases, fatty alcohol esterases, terpene alcoholases, and terpene alcohol esterases, i.e. the enzymes which break down the ester linkages in the wood resin. It is, moreover, advantageous if they produce enzyme systems which break down the carbon chains of the resin compounds by oxidation. The micro-organisms should preferably also be sporulating since the spores can then easily be added to the wood chips in the form of an aqueous dispersion, but if the micro-organisms are constituted of non-sporulating fungi it is naturally also possible to break down the same prior to adding said organisms. The said micro-organisms should suitably also be resistant to low temperatures, to enable them to be used when storing wood chips in winter.

Examples of suitable resin decomposing micro-organisms which do not produce active cellulases on wood to any appreciable degree, and which inhibit cellulose and hemicellulose decomposing micro-organisms by developing such a high temperature in the pile of wood chips that said micro-organisms cannot propagate are, i.a.:

connected to the duct 2. The slurry is injected into the chips by means of nozzles 9, while they are being transported to the storage site along the duct 2. The following examples illustrate the invention:

(1) Humicola grisea var. thermoidea, characterized by Example 1 i gzg figzf jg ch y Bunce Wax from spruce wood was isolated by extraction of (3) Humicola insole; by C0Oney EmerSOn spruce chips with ethanol-benzene (1:2), evaporation, Humicola [an Ch b GrifiomMaublano extraction w1th ethyl ether and agitation of the ether- Bunce g y soluble portion with KOH, whereby the acid substances Talarom Ces d anti ch b Grifion Maublanc present in the resin were converted into their water- Coone infers p y soluble potassium salts, which were separated. The neu- Thefmoascus aumnfiacus ch b Miehe tral portion was subjected to chromatography on a silica- Mucor minus ch b i y gel column and the eluate containing wood waxes was Mucor 5 'y g y Emerson mixed, subsequent to evaporation, with a nutritional salt (9) Malbranclzea pulchella, Saccardo-Penzig var. sulf- Sohmon contammg the followmg mgwdlents' M0 urea, ch. by Miehe KH 56 2 4 (l0) Myrzococcum albomyces, ch. by Cooney-Emerson KHPO 150 (11) Sporothrichu'm thermophile, ch. by Apinis NH 6 500 (12) Allescheria terrestris, ch. by Apinis M 4 3 (13) Torula thermophzla, ch. by Cooney-Emerson Fe citrate 20 (14) Chaetomium thermophile, ch. by La Touche Yeast y 1O (15) Stilbella thermophile, ch. by Fergus Distilled 'ibbgg (16) Aspergillus fumigatus, ch. by Fresenius Examples of suitable resin decomposing g D The concentration of resin in the nutritional salt solunisms which do not produce active cellulases on wood g a sgg ggg g g ig 33 :5 is; 5 8; 3; 2335;: and which inhibit cellulose and hemicellulose decomposf g ing micro-organisms by antibiosis effect are i.a. the folo Vain-ms mlsxo'olgamsms accor o e mven Ion lowing were in ected mto or on the emulsions, under the condltions which can be seen from Table 1, after which (17) Trichoderma lignorum, ch. by Tode-Harz decomposition of the wax was determined. These micro- (18) Gliocladium roseum, ch. by Link-Thom organisms were isolated from wood samples taken from (19) Gliocladium deliquescens, ch. by Sopp wood chip piles by sterile transfer to agar plate cultures, (20) Pcnicillium funicolosum, ch. by Thom wherefrom the separate micro-organisms were separated (21) Penicillium roqueforti, ch. by Thom from the various colonies obtained and re-cultivated on (22) Penicillium rubrum, oh. by Stoll agar plates. The spores from the thus re-cultivated micro- (23) Gliocladium viride organisms were isolated by flushing with water.

TABLE 1 Temp., Agitated Stationary Microorganism 0. Days culture culture Wax decomposition Penicilljlmtvt 25 4 No trace of wax.

TU M8 0T 1. P;n z ctlli1ZLm 25 8 Certain wig: de- U/IZZCU osum. CO 0 l 1011. Bfisggilfimys 16 No 3 231501 wax. Trtchodermiz 25 8 Certain wax delzgnorum. composition. Rhizohpus 5O 4 Wax gonsurrtilption (ZTT 221M}. Dactylomz/ces 4 l of thermophilus. Penicillium 13-76-1 50 4 Wax decomposi- Sporotrichum 50 4 l ti and growth.

thermophile. u Allescheria 50 4 Do.

terrestris.

Other suitable fungi for use according to the invention, associated with any of said groups are the following:

(24) Penicillium, our designation B76-1, a new species being characterized (25) Dactylomyses thermophilus (26) Rhizopus arrhizus (27) Byssochlamys, our designation M 243 II, a new species being characterized (28) Penicillium, our designation P 116I, a new species being characterized A suitable apparatus for applying the method is shown diagrammatically in the figure. A plant for building a pile of wood chips is shown in the form of a blower machine 1, a pneumatic conveying duct 2, a chip hopper 3 with a discharge device (cell feeder) 4 connected to the duct 2. A tank 5 intended to contain a slurry of micro-organisms is connected via conduits 8 with a pump 6 and an overflow valve 7 in connection with nozzles 9 in the duct 2 on both sides of the junction where the cell feeder 4 is The test shows that the above micro-organisms, which were all isolated from piles of wood chips, are capable of developing, using wax present in the nutritional solution as a source of carbon. The decomposition of wax which occurs when storing wood chips can thus be attributed to certain of the micro-organisms which are developed in the pile of chips.

Example 2 temperature organism. The samples were then extracted with ethyl alcohol and benzene in the ratio of 1:2, after which the extract was evaporated and treated with ethyl ether. The ether-soluble portion was made up of the resin remaining in the wood. This was agitated with KOH,

were placed at a level of 5 meters above the ground. At the same time 5 sacks containing untreated wood chips removed direct from the storage container were placed at the same level, for checking purposes.

The Weight of the chips and the bulk density, extract acid substances (free fatty acids and resin acids) present 5 content DKM, lignin content, pentosan content, carboin the resin being converted into water-soluble potassium hydrate composition and solids content of the chips in salts, which were separated. The remaining ether solution each sack were determined beforehand. Two months later was examined quantitatively and qualitatively chromatothe pile was broken down and the sacks removed, after graphically on its content of hydrocarbons, waxes, tri- 10 which the above analyses were repeated. Temperature glycerides, higher alcohols and other substances. At the development in the sacks during storage was determined same time completely untreated chips were analysed, said at various levels. Each chip portion placed in the pile chips having been stored under corresponding conditions was test cooked according to the sulphate method under (control). The result can be seen from Table 2. standard conditions to 35 cp. viscosity, after which the TABLE 2 Rhizpous Byssochlamys Definition Control arrhizus M 243-11 Storage time, days 0 17 17 30 60 60 30 60 17 30 60 Storage temp., C 50 35 50 30 50 50 50 50 50 50 EtOH/Benzcne (112) extract 3.248 2.645 2.676 2.616 1.735 3.395 2.087 1.721 1.679 2.499 2.114 1.507 Insoluble in ether, ercent of wood 1. 72 1. 068 1.139 1. 728 1. 050 2. 422 1. 232 1. 089 1.064 1. 304 1. 291 0. 927 Soluble in ether, percent of wood 1. 53 1. 139 1. 006 0.888 0.685 0. 974 0. 856 0. 633 0.616 0. 829 0. 823 0.580 Neutral substances, percent of wood- 1. 024 0.541 0. 474 0. 405 0. 355 0. 380 0. 311 0. 344 0. 295 0. 472 0.424 0. 311 Hydrocarbons, percent of wood. 0. 045 0. 018 0. 023 0. 018 0. 015 0.015 0. 013 0. 011 0. 012 0. 025 0. 016 0. 011 Waxes, percent of wood 0. 177 0.115 0. 110 0.083 0.073 0.059 0.029 0.045 0.026 0.131 0.078 0.037 Triglycerides, percent of wood. 0.532 0. 106 0. 061 0.068 0. 020 0. 092 0.062 0. 059 0.075 Higher, alcohols, percent otwood. 0.116 0.139 0.146 0.151 0.135 0.109 0.171 0.134 0.171 0.126 0.143 0.167 Remainder, percent ofwood 0.150 0.164 0.135 0. 085 0.114 0.103 0.096 0.091 0. 087 0.131 0.112 0.097 Free acids, ercent orwood 0. 514 0. 598 0. 532 0. 483 0.330 0. 594 0. 545 0. 289 0. 321 0.357 0.399 0. 259 Free fatty acids, percent of wood. 0. 086 0. 146 0.083 0.064 0. 091 Resin acids, percent of wood 320 0.382 0. 319 0. 19 0. 137 Loss in weight percent of original wood, probable value 0. 95 2. 29 1. 23 4. 77 4. 9 4. 7 4. 58 2. 6 1. 23 1. 3. 8 Lignirl acc. to Klaso 27. 8 28.1 27.07 27.33 28. 33 28.12 27.43 28. 69 28. 5 27.28 29. 49

Penicillium Gliorladium Penicillium Trichoderma. Penicillium Aspergillus Definition P vin'de rubmm lignorum roqueforti fumigatus Storage time, days 30 60 30 60 30 60 30 60 17 30 60 30 60 Storage tern C 35 35 35 35 35 35 35 35 35 35 35 50 50 EtOH/Benzene (1:2) extract 2.751 1.865 2.070 1.900 1.972 2.208 2.345 1.907 3.112 2.084 2.012 2.316 1.734 Insoluble in ether, percent of wood 1.890 0. 921 1.139 1. 095 1.117 1. 457 1. 258 1. 091 1. 998 1.188 1. 082 1. 435 1. 094 Soluble in ether, percent of wood 0.861 0.944 0.932 0.806 0.855 0.751 1.087 0.816 1.114 0.896 0.930 0.881 0.639 Neutral substances, percent of wood.. 0.399 0.487 0.437 0.391 0.448 0.397 0.493 0.375 0.509 0.464 0.390 0.489 0.235 Hydrocarbons, percent or wood 0.011 0.017 0.012 0.013 0.016 0. 017 0.047 0.010 0.018 0. 011 0.013 0.019 0. 017 Waxes, percent or wood 0.074 0.090 0.056 0.059 0.084 0.065 0. 087 0.058 0.131 0.071 0.063 0.102 0.022 Triglycerides, percent oiwood 0.085 0.083 0.089 0. 071 0.083 0.080 0. 087 0.078 0.107 0. 085 0.083 0.069 Higher alcohols, percent of wood. 0.110 0.162 0.109 0.145 0.130 0.127 0.166 0.119 0.147 0.141 0.144 0.155 0.131 Remainder, percent of wood... 0. 156 0.088 0. 144 0.065 Free acids, percent of wood 0 0. 432 0.540 0. 392 0. 404 Free fatty acids, percent of wood. 0. 129 0.213 Resin acids, percent of Wood 0.329 0. 159 Loss in weight percent of origlnal wood,

probable value 5. 25 7. 4 3. 07 1. 5 2. 46 2. 6 0. 67 2. 7 1. 51 0. 2. 9 0. 89 0. 9

Lignin acc. to Klason As can be seen from the tables the decomposition of resin is considerably improved when applying the invention, in comparison with conventional methods of storing wood chips (compare soluble in ether in Table 2, line 3). For example the micro-organism Byssochlamys M 243-11 gives an approximately 20% improvement in this respect. The amount of residual wax with the organisms used in the example was substantially unchanged whilst the loss of wood was greatly reduced.

Example 3 A pile of chips 25 x 25 x 10 meters in dimension (2000 m?) was built by means of the apparatus shown in the figure, said apparatus comprising a blower machine 1, a pneumatic conveying duct 2, a supply container for wood chips 3, in the lower portion of which was arranged a discharge device sealing against the supply container (cell feeder) 4. From a tank 5 with a capacity of 200 litres and containing 15.10 slurried spores of the microorganisms Rhizopus arrhizus (high-temperature fungus) and Gliocladium viride (low-temperature fungus), the slurry was pumped by pump 6 via the over-flow valve 7 through the line 8 to the nozzles 9 arranged on both sides of the cell feeder into the pneumatic transport duct 2 at the same time as wood chips were fed into the said pneumatic transport duct. The slurry was delivered in an amount of 53 litres per hour whilst the chips were fed in at a speed of approximately m. /hour. To determine the loss of wood, five sample sacks of wood chips obtained pulp was bleached according to the following 1 Times kappa.

Corresponding cooks and bleaches were carried out subsequent to storing the chips according to the invention. It was shown that the chips treated with microorganisms according to the invention presented a total loss in weight of 1.3% whereas the untreated chips showed a loss in weight of 2.2%. The weight loss of the untreated portions must be considered low since the normal value for losses in weight can lie between 3-4%. The reasons why the values were so low in this case was due to the fact that all the surrounding chips had been treated and thus protected the untreated chip portions from deleterious micro-organisms.

On analysis of micro-flora present in the pile of Wood chips it was seen that in the outer portions of the pile, where the low-temperature fungus Gliocladium viride had been active, the protective eflfect by anti-biosis had been complete. The amount of rot fungi in the inner portion of the pile was very slight. The total protective effect had thus been very good. The extract content (DKM) of chips treated according to the invention, and unbleached and bleached sulphite pulp produced from said chips, was 0.70, 0.85 and 0.13%. Corresponding values for untreated chips and unbleached and bleached sulphite pulp produced from said chips was 0.90, 1.30 and 0.20%.

Concerning other analysis values no difference could be noted with pulp produced from chips treated according to the invention or with pulps produced from chips stored in the conventional manner.

We claim:

1. A process for storing wood chips under conditions such that resinous components present in the chips are subjected to enzymatic hydrolysis induced by fungi added to the chips while minimizing microbial action on cellulose and hemicellulose in the wood, which comprises adding to the wood chips a fungus or fungi capable of elaborating esterases which attack ester linkages in the wood resinous components and which either completely or partly lack the ability of elaborating active cellulases, selected from the group consisting of:

(l) fungi which inhibit celluloseand hemicellulosedecomposing microorganisms by developing a high temperature in the wood chips inhibiting propagation of such micro-organisms, selected from Humicola grisea var. thermoideal, characterized by Cooney-Emerson Humicola stellata, characterized by Bunce H umzcola insolens, characterized by Cooney-Emerson Humicola languin'osa, characterized by Griffon- Maublanc-Bunce T alaromyes duponti, characterized by Grifion- Maublanc-Cooney-Emerson Thermoascus aurantiacus, characterized by Miehe Mucor pusillus, characterized by Lindt Mucor miehi, characterized by Cooney-Emerson Malbranchea pulchella, Saccardo-Penzig var. sulfur-ea, characterized by Miehe Myriococcwm albomyces, characterized by Cooney-Emerson Sporothrichum Apinis Allescheria terrestris, characterized by Apinis thermophile, characterized 'by 8 Torula thermophile', characterized by Cooney- Emerson Chaetomium zhermophile, characterized by La Touche Stilbella zhermophile, characterized by Fergus Aspergz'llus fumigatus, characterized by Fresenius (2) fungi which inhibit celluloseand hemicellulosedecomposing micro-organisms by antibiotic effect, selected from:

Trichoderma Lingnorum, characterized by Tode- Harz Gliocladium roseum, characterized by Link-Thom Gliocladium deliquescens, characterized by Sopp Penicillium funicolosum, characterized by Thom Penicillium roqueforti, characterized by Thom Penicillium rubrum, characterized by Stoll Gliocladium viride and fungi selected from:

D'actylomyses thermophilus Rhizopus arrhizus and then storing the wood chips in a pile under conditions favorable to the growth of the added fungus or fungi.

2. A process in accordance with claim 1 in which the fungi are added to the wood chips in the form of an aqueous dispersion of the spores thereof.

3. A process in accordance with claim 1 in which the fungus is especially selected to propagate at low winter temperatures.

4. A process in accordance with claim 1 in which the fungi are applied to the chips in the form of an aqueous dispersion which is sprayed over the chips.

References Cited UNITED STATES PATENTS 1,633,594 6/1927 Lathrop -8 X 2,193,493 3/1940 Ritter 195-10 2,871,163 1/1959' Turnbull 195-8 3,401,085 9/1968 Croon l62-1 X HOWARD R. CAINE, Primary Examiner US. Cl. X.R. 

